Vascular endothelial cell dysfunction, mediated by changes in redox cell signaling, plays an important role in the etiology of a number of chronic diseases such as atherosclerosis and hypertension. Nitric oxide (NO) has a critical role in vascular physiology generated at low levels by eNOS and the pathophysiology of a wide array of cardiovascular disorders when formed in conjunction with reactive oxygen species or at high levels by iNOS. An important target for NO in the endothelium that has received little attention in the context of redox cell signaling is the mitochondrion. Mitochondrial dysfunction can occur in the endothelium on exposure to reactive oxygen and nitrogen species (ROS/RNS) and could contribute to the development of atherosclerosis through mechanisms that have yet to be defined. We have recently proposed that the "NOcytochrome c oxidase signaling pathway" is a major target for NO dependent effects in endothelial cells through controlling formation of superoxide and hydrogen peroxide by the organelle. The mediators between the mitochondrion and cytosol resulting in redox signaling are proposed to be electrophilic lipid oxidation products including the cyclopentanones and aldehydes such as 4-hydroxynonenal. In support of this concept preliminary data demonstrates the potential of these lipid species to activate signal transduction pathways including the Keapl/nrf2 system. These data led to the hypothesis that NO-dependent control of mitochondrial function contributes to redox cell signaling through the transduction of nitrosative cell signaling to the controlled formation of oxidative stimuli. This hypothesis will be investigated using a variety of molecular and cellular approaches including the proteomics analysis of endothelial cells to identify ROS/RNS dependent post translational modification of proteins that contribute to redox cell signaling. The proposal builds upon previous experimental findings through pursuit of the following specific Aims; 1: Determine the effect of mitochondrial dysfunction ROS formation from the organetle in endothelial cells. Specific Aim 2: Determine the impact of mitochondrial ROS formation on the post-translational modification of endothelial cell proteins during oxidative stress. Specific Aim 3: Determine the effect of controlling mitochondrial ROS formation on redox cell signaling. The insights gained by the accomplishment of these specific aims will help in understanding the role of NO in modulation of mitochondrial function in the vasculature. This would then enable the design of therapeutic strategies in treatment of chronic vascular diseases such as atherosclerosis. [unreadable] [unreadable]